Evolution of EPSPS double mutation imparting glyphosate resistance in wild poinsettia (Euphorbia heterophylla L.).

The evolution of glyphosate resistance (GR) in weeds is an increasing problem. Glyphosate has been used intensively on wild poinsettia (Euphorbia heterophylla L.) populations for at least 20 years in GR crops within South America. We investigated the GR mechanisms in a wild poinsettia population from a soybean field in southern Brazil. The GR population required higher glyphosate doses to achieve 50% control (LD50) and 50% dry mass reduction (MR50) compared to a glyphosate susceptible (GS) population. The ratio between the LD50 and MR50 of GR and GS resulted in resistance factors (RF) of 6.9-fold and 6.1-fold, respectively. Shikimate accumulated 6.7 times more in GS than in GR when leaf-discs were incubated with increasing glyphosate concentrations. No differences were found between GR and GS regarding non-target-site mechanisms. Neither population metabolized glyphosate to significant levels following treatment with 850 g ha-1 glyphosate. Similar levels of 14C-glyphosate uptake and translocation were observed between the two populations. No differences in EPSPS expression were found between GS and GR. Two target site mutations were found in all EPSPS alleles of homozygous resistant plants: Thr102Ile + Pro106Thr (TIPT-mutation). Heterozygous individuals harbored both alleles, wild-type and TIPT. Half of GR individuals were heterozygous, suggesting that resistance is still evolving in the population. A genotyping assay was developed based on the Pro106Thr mutation, demonstrating high efficiency to identify homozygous, heterozygous or wild-type EPSPS sequences across different plants. This is the first report of glyphosate-resistant wild-poinsettia harboring an EPSPS double mutation (TIPT) in the same plant.

Chemical Characterization and Phytotoxic Effects of the Aerial Parts of Ruzigrass (Urochloa ruziziensis).

Studies of the phytotoxic effects between plants can be a crucial tool in the discovery of innovative compounds with herbicide potential. In this sense, we can highlight ruzigrass (Urochloa ruziziensis), which is traditionally used in the crop rotation system in order to reduce weed emergence. The aim of this work was to characterize the secondary metabolites of ruzigrass and to evaluate its phytotoxic effects. In total, eight compounds were isolated: friedelin, oleanolic acid, α-amyrin, 1-dehydrodiosgenone, sitosterol and stigmasterol glycosides, tricin and p-coumaric acid. Phytotoxic effects of the crude methanolic extract and fractions of ruzigrass were assessed using germination rate, initial seedling growth, and biomass of Bidens pilosa, Euphorbia heterophylla and Ipomoea grandifolia. Chemometric analysis discriminated the weed species into three groups, and B. pilosa was the most affected by fractions of ruzigrass. The phytotoxic activities of 1-dehydrodiosgenone, tricin, and p-coumaric acid are also reported, and p-coumaric acid and 1-dehydrodiosgenone were active against B. pilosa.

MicroRNAome Profile of Euphorbia kansui in Response to Methyl Jasmonate.

miRNAs play vital regulatory roles in different plant developmental stages and in plant response to biotic and abiotic stresses. However, information is limited on the miRNA regulatory mechanism to methyl jasmonate (MeJA). In this study, we used the microRNAome profile to illustrate the relevant regulatory mechanisms of Euphorbia kansui in response to methyl jasmonate (MeJA) through Illumina RNA-Seq. As a result, we identified 875 miRNAs corresponding to 11,277 target mRNAs, among them, 168 known miRNA families representing 6019 target mRNAs sequences were obtained. 452 miRNA-mRNA pairs presented an anti-correlationship (Cor < -0.50 and p-value of correlation ≤ 0.05). The miRNA with a fold change ≥ 2 and a p (p-Value) < 0.05 in pairwise comparison were identified as significant differentially expressed miRNAs (DEMs). The DEMs in MeJA treatment of 0, 24, 36 and 48 h were compared by using Short Time Expression Miner (STEM) cluster and 4 significant gene profiles (p-value ≤ 0.02) were identified. Through the kyoto encyclopedia of genes and genomes (KEGG) pathway and gene ontology (GO) enrichment analysis on all miRNA targets, we identified 33 mRNAs in terpenoid biosynthesis, which were regulated by miRNAs under MeJA treatment, so the miRNA maybe involved in the response of E. kansui plant to exogenous MeJA and the results would provide very useful information on illustrating the regulatory mechanism of E. kansui and also provide an overall view of the miRNAs response to MeJA stress of a non-model plant.

Foliar Glyphosate Treatment Alters Transcript and Hormone Profiles in Crown Buds of Leafy Spurge and Induces Dwarfed and Bushy Phenotypes throughout its Perennial Lifecycle.

Leafy spurge ( L.) is an invasive weed of North America and its perennial nature attributed to underground adventitious buds (UABs) that undergo seasonal cycles of para-, endo-, and ecodormancy. Recommended rates of glyphosate (∼1 kg ha) destroy aboveground shoots but plants still regenerate vegetatively; therefore, it is considered glyphosate-tolerant. However, foliar application of glyphosate at higher rates (2.2-6.7 kg ha) causes sublethal effects that induce UABs to produce stunted, bushy phenotypes. We investigated the effects of glyphosate treatment (±2.24 kg ha) on vegetative growth, phytohormone, and transcript profiles in UABs under controlled environments during one simulated seasonal cycle. Because shoots derived from UABs of foliar glyphosate-treated plants produced stunted, bushy phenotypes, we could not directly determine if these UABs transitioned through seasonally induced endo- and ecodormancy. However, transcript abundance for leafy spurge dormancy marker genes and principal component analyses suggested that UABs of foliar glyphosate-treated plants transitioned through endo- and ecodormancy. Glyphosate treatment increased shikimate abundance in UABs 7 d after treatment; however, the abundance of shikimate gradually decreased as UABs transitioned through endo- and ecodormancy. The dissipation of shikimate over time suggests that glyphosate's target site was no longer affected, but these changes did not reverse the altered phenotypes observed from UABs of foliar glyphosate-treated leafy spurge. Transcript profiles further indicated that foliar glyphosate treatment significantly affected phytohormone biosynthesis and signaling, particularly auxin transport; gibberellic acid, abscisic acid and jasmonic acid biosynthesis; ethylene responses; and detoxification and cell cycle processes in UABs. These results correlated well with the available phytohormone profiles and altered phenotypes.

Effect of endophytic Bacillus cereus ERBP inoculation into non-native host: Potentials and challenges for airborne formaldehyde removal.

Phytoremediation could be a cost-effective, environmentally friendly approach for the treatment of indoor air. However, some drawbacks still dispute the expediency of phytotechnology. Our objectives were to investigate the competency of plant growth-promoting (PGP) endophytic Bacillus cereus ERBP (endophyte root blue pea), isolated from the root of Clitoria ternatea, to colonize and stabilize within Zamioculcas zamiifolia and Euphorbia milii as non-native hosts without causing any disease or stress symptoms. Moreover, the impact of B. cereus ERBP on the natural shoot endophytic community and for the airborne formaldehyde removal capability of non-native hosts was assessed. Non-native Z. zamiifolia was effectively inoculated with B. cereus ERBP through soil as the most efficient method of endophyte inoculation. Denaturing gradient gel electrophoresis profiling of the shoot endophytic community verified the colonization and stability of B. cereus ERBP within its non-native host during a 20-d fumigation period without interfering with the natural shoot endophytic diversity of Z. zamiifolia. B. cereus ERBP conferred full protection to its non-native host against formaldehyde phytotoxicity and enhanced airborne formaldehyde removal of Z. zamiifolia whereas non-inoculated plants suffered from formaldehyde phytotoxicity because their natural shoot endophytic community was detrimentally affected by formaldehyde. In contrast, B. cereus ERBP inoculation into non-native E. milii deteriorated airborne formaldehyde removal of the non-native host (compared to a non-inoculated one) as B. cereus ERBP interfered with natural shoot endophytic community of E. milii, which caused stress symptoms and stimulated ethylene biosynthesis. Non-native host inoculation with PGP B. cereus ERBP could bear potentials and challenges for airborne formaldehyde removal.

Glyphosate's impact on vegetative growth in leafy spurge identifies molecular processes and hormone cross-talk associated with increased branching.

BACKGROUND: Leafy spurge (Euphorbia esula) is a perennial weed that is considered glyphosate tolerant, which is partially attributed to escape through establishment of new vegetative shoots from an abundance of underground adventitious buds. Leafy spurge plants treated with sub-lethal concentrations of foliar-applied glyphosate produce new vegetative shoots with reduced main stem elongation and increased branching. Processes associated with the glyphosate-induced phenotype were determined by RNAseq using aerial shoots derived from crown buds of glyphosate-treated and -untreated plants. Comparison between transcript abundance and accumulation of shikimate or phytohormones (abscisic acid, auxin, cytokinins, and gibberellins) from these same samples was also done to reveal correlations. RESULTS: Transcriptome assembly and analyses confirmed differential abundance among 12,918 transcripts (FDR ≤ 0.05) and highlighted numerous processes associated with shoot apical meristem maintenance and stem growth, which is consistent with the increased number of actively growing meristems in response to glyphosate. Foliar applied glyphosate increased shikimate abundance in crown buds prior to decapitation of aboveground shoots, which induces growth from these buds, indicating that 5-enolpyruvylshikimate 3-phosphate (EPSPS) the target site of glyphosate was inhibited. However, abundance of shikimate was similar in a subsequent generation of aerial shoots derived from crown buds of treated and untreated plants, suggesting EPSPS is no longer inhibited or abundance of shikimate initially observed in crown buds dissipated over time. Overall, auxins, gibberellins (precursors and catabolites of bioactive gibberellins), and cytokinins (precursors and bioactive cytokinins) were more abundant in the aboveground shoots derived from glyphosate-treated plants. CONCLUSION: Based on the overall data, we propose that the glyphosate-induced phenotype resulted from complex interactions involving shoot apical meristem maintenance, hormone biosynthesis and signaling (auxin, cytokinins, gibberellins, and strigolactones), cellular transport, and detoxification mechanisms.

Micropropagation of poinsettia by organogenesis.

Poinsettia (Euphorbia pulcherrima) is one of the most popular ornamental pot plants. Conventional propagation is by cuttings, generally focused on a period prior to the most intensive time of sales. Rapid multiplication of elite clones, the production of pathogen-free plants and more rapid introduction of novel cultivars (cvs.) with desirable traits, represent important driving forces in the poinsettia industry. In recent years, different strategies have been adopted to micropropagate poinsettia, which could assist breeders to meet consumer demands. The development of reliable in vitro regeneration procedures is likely to play a crucial role in future production systems. Stem nodal explants cultured on semi-solid MS-based medium supplemented with benzylaminopurine (BAP) and naphthalene acetic acid (NAA) develop shoots from adventitious/axillary buds after 7 weeks of culture. Rooting of in vitro regenerated shoots is achieved in semi-solid MS-based medium containing the auxin indole-3-acetic acid (IAA). Four to six weeks after transfer to root-inducing medium, regenerated plants can be transferred to compost and acclimatized in the glasshouse. Direct shoot regeneration from cultured explants is important to minimize somaclonal variation in regenerated plants.

The phytotoxic effect of exogenous ethanol on Euphorbia heterophylla L.

This study investigated the effects of exogenously applied ethanol on Euphorbia heterophylla L., a troublesome weed in field and plantation crops. Ethanol at concentrations ranging from 0.25 to 1.5% caused a dose-dependent inhibition of germination and growth of E. heterophylla. Measurements of respiratory activity and alcohol dehydrogenase (E.C. 1.1.1.1) activity during seed imbibition and initial seedling growth revealed that ethanol induces a prolongation of hypoxic conditions in the growing tissues. In isolated mitochondria, ethanol inhibited the respiration coupled to ADP phosphorylation, an action that probably contributed to modifications observed in the respiratory activity of embryos. A comparison of the effects of methanol, ethanol, propanol and acetaldehyde on germination and growth of E. heterophylla indicates that alcohol dehydrogenase activity is required for the observed effects, with the conversion of ethanol to acetaldehyde playing a role in the ethanol-induced injuries.

Control effort exacerbates invasive-species problem.

Ecosystem managers face a difficult decision when managing invasive species. If they use aggressive practices to reduce invader abundances, they will likely reduce invaders' competitive impacts on natives. But it is often difficult or impossible to reduce invaders without damaging natives. So a critical question becomes: Which is worse for native biota, invaders or things done to control invaders? We attempted to answer this question for a common scenario. We studied several grassland natives exhibiting long-term coexistence with an invader and asked how aggressive management (herbicide use) affected the natives. Whether or not grazing was excluded, one-time herbicide use made two native forbs exceedingly rare for our entire 16-year study period. Herbicide also made several other native forbs rare, but only when grazing was excluded, and there is evidence that the dominant invader became more abundant in response to the decreases in native-forb abundances. Throughout the world, terrestrial and aquatic ecosystems are receiving herbicide applications for exotic-species control. Some of the applications are doubtless warranted because they target small invader patches or larger areas with virtually no remaining natives. However, other herbicide applications occur where large native populations occur, and our data suggest that these applications can be ill advised. Our cautionary tale is told using an herbicide-treated grassland, but our results should be considered wherever invasive-species management damages native species.

Synthesis of new phytogrowth-inhibitory substituted aryl-p-benzoquinones.

Reaction of [(2-alkyloxy)methyl]-1,4-dimethoxybenzene 10 (alkyl=butyl, hexyl, decyl, tridecyl, tetradecyl, hexadecyl, and octadecyl) with ceric ammonium nitrate in order to produce p-benzoquinones (=cyclohexa-2,5-diene-1,4-diones) afforded 5-[(alkyloxy)methyl]-2-(4-formyl-2,5-dimethoxyphenyl)benzo-1,4-quinones 12a-12g in yields that varied from 46 to 97%, accompanied by 2-[(alkyloxy)methyl]benzo-1,4-quinones 11a-11g in only small quantities (< or =5%). These quinones resemble the natural phytotoxic compound sorgoleone, found in Sorghum bicolor. This reaction exemplifies a general procedure for the synthesis of novel aryl-substituted p-benzoquinones. The selective effects of compounds 12a-12g, at the concentration of 5.5 ppm, on the growth of Cucumis sativus, Sorghum bicolor, Euphorbia heterophylla, and Ipomoea grandifolia were evaluated. All compounds caused some inhibition upon the aerial parts and root growth of the tested plants. The most active compound, 2-(4-formyl-2,5-dimethoxyphenyl)-5-[(tridecyloxy)methyl]-benzo-1,4-quinone (12d), caused between 3 and 18%, and 12 and 29% inhibition on the roots and aerial parts development of Cucumis sativus and Sorghum bicolor, respectively, and between 77 and 85%, and 34 and 52% inhibition on the roots and aerial parts growth of Euphorbia heterophylla and Ipomoea grandifolia, respectively.

Synthesis of new aliphatic and aromatic phytotoxic derivatives of 2alpha,4alpha-dimethyl-8-oxabicyclo[3.2.1]oct-6-en-3-one.

Several new compounds with potential herbicidal activity were synthesized from 2alpha,4alpha-dimethyl-6,7-exo-isopropylidenedioxy-8-oxabicyclo[3.2.1]octan-3-one (4). Seven aromatic alcohols were prepared by reaction of (4) with aryllithium reagents, where the aryl groups were 4-ethoxyphenyl (5, 70% yield), 4-ethylphenyl (6, 82% yield), 4-butylphenyl (7, 78% yield), 4-tert-butylphenyl (8, 81% yield), 2,4-dimethoxyphenyl (9, 75% yield), 2-ethylphenyl (10, 12% yield) and para-(4-bromophenoxy)phenyl (11, 24% yield). Reaction of the acetonide (4) with Grignard reagents formed also four aliphatic alcohols where the alkyl groups are ethyl (13, 78%), butyl (14, 85%), hexyl (15, 81%) and octyl (25, 92%). The alcohols (5), (6), (7), (8), (13), (14), (15) and (25) were reacted with thionyl chloride in pyridine, forming their respective alkenes (17, 76%), (18, 74%), (19, 83%), (20, 73%), (22, 78%), (26, 62%), (23, 77%) and (24, 66%). The effect of these compounds, at the concentration of 5.5 microg g(-1), on the development of radicle and aerial parts of Sorghum bicolor (L) Moench, Euphorbia heterophylla L, Brachiaria decumbens and Desmodium tortuosum DC was evaluated.

The Ecological Areawide Management (TEAM) of leafy spurge program of the United States Department of Agriculture-Agricultural Research Service.

The Ecological Areawide Management (TEAM) of Leafy Spurge program was developed to focus research and control efforts on a single weed, leafy spurge, and demonstrate the effectiveness of a coordinated, biologically based, integrated pest management program (IPM). This was accomplished through partnerships and teamwork that clearly demonstrated the advantages of the biologically based IPM approach. However, the success of regional weed control programs horizontally across several states and provinces also requires a vertical integration of several sectors of society. Awareness and education are the essential elements of vertical integration. Therefore, a substantial effort was made to produce a wide variety of information products specifically designed to educate different segments of society. During its tenure, land managers and agency decision makers have seen the potential of using the TEAM approach to accelerate the regional control of leafy spurge. The example set by the TEAM organization and participants is viewed as a model for future weed-control efforts.

Cross resistance to ALS-inhibiting herbicides in Euphorbia heterophylla L. biotypes resistant to imazethapyr.

The effect of weeds on the diminution of agricultural production is considered to be between 30 and 50%. Imazethapyr is a herbicide which acts on the enzyme ALS, the first common enzyme in the biosynthetic pathway of valine, leucine and isoleucine. Euphorbia heterophylla is a common species in soybean fields in Brazil and different populations resistant to herbicides of the sulfonylurea and imidazolinone groups are currently being reported. The objectives of this work were: to determine the resistance of three different biotypes named RI, RII and RII to the herbicide imazethapyr: to establish the level of resistance to this herbicide, and to evaluate the behaviour of the biotypes to other herbicides with a similar mode of action. The response of plants treated with imazethapyr confirmed the existence of resistance in the biotypes denominated as RI, RII and RIII. The imazethapyr concentration required to reduce the activity of the enzyme ALS by 50% (I50) should be greater than 500 microM for biotypes RI, RII and RIII and 39.47 microM for the susceptible biotype. Applications made in the greenhouse at field doses of ALS inhibiting herbicides showed cross resistance in the case of imazamox and imazamethabenz but not in the cases of the other ALS inhibitor herbicides used. ALS activity results demonstrated that the imazethaphyr resistance in different biotypes of E. heterophylla is target site mediated.